Monday, July 25, 2011

Kevin Ahern (www.davincipress.com/metabmelodies.html) is a professor at the Oregon State University who have created several musics about biochemistry for his students. Here is the lyrics of the music about gluconeogenesis (based on the song Supercalifragilisticexpialidocious) and the link for the download of the .mp3 file.

Sunday, July 24, 2011

This reaction is clearly favored in the forward direction. In situations of hypoxia (intense muscular effort, for example), or absence of mitochondria, the cell is unable to regenerate NAD+ from NADH through the respiratory chain. It does so using the conversion of pyruvate to lactate, which consumes NADH and releases NAD+ release. It should be noted that the NAD+ is essential for glycolysis to continue to occur, thereby to obtain energy through the catabolism of sugars. Each molecule of pyruvate converted to lactate regenerates a molecule of NAD+. The lactate formed is sent through the bloodstream to the liver where it is converted to glucose in gluconeogenesis. The question that arises is: "So if you can recycle lactate, converting it back into glucose, why is the liver that has to do this and not the muscle, since it is the muscle that produces lactate? If so, the muscle could use directly the product of fermentation to restore the levels of metabolic fuel." In fact, at a first glance it may make sense to think in this way. However, the synthesis of glucose through gluconeogenesis is very expensive, in terms of energy, so that after an intense physical effort, it did not make sense that the muscle has to spend additional energy to synthesize glucose. Thus, the recovery of an intense effort includes not only the restoration of ATP levels in muscle but also an extra consumption of oxygen in the liver, necessary for the synthesis of ATP to be used in gluconeogenesis from lactate. In other words, after muscular efforts, is the liver that has to use lactate, allowing a faster and more efficient muscle recovery. This process is called the Cori cycle.

Dring anaerobic work the concentration of lactate in the muscle fibers can increase about 30 times and it is a commonplace to say that it is this accumulation of lactate ion which causes fatigue. However, the experimental evidence shows that although the concentration of lactate is directly related to the degree of fatigue it does not interfere with the the muscle contractile activity. Fatigue, muscle pain and cramping experienced after an intense physical effort are the result of an acidification caused by lactic acid in muscle (the pH can drop from 7 to 6.5 !!!). The pKa of lactic acid is about 4, which causes that at the cell pH (≈ 7) or plasma (≈ 7.4) occurs the dissociation of lactic acid to lactate + H+. This accumulation of H+ will interfere with the contractile capacity of muscle fibers and will also invade the synaptic cleft. Thus, the inability of the neuromuscular junction in relaying the nerve impulses to muscle fibers is due probably to a lower release of the chemical transmitter acetylcholine by nerve endings, due to acidification of the interstitial fluid and alteration of protein structures (acetylcholine receptors) by the action of H+. This system provides energy for physical activities that result in fatigue after about 60-120 seconds. It is therefore the primary metabolic process associated with activities such as running up to 400-800 m, swimming events of 100-200m, and also provides energy for high intensity moments in football, basketball, volleyball, tennis, among others. The common denominator of these activities is the support of high-intensity efforts lasting 1-2 minutes. Even the best trained athletes are unable to sprint for more than a minute. A highly competitive athlete needs about 30 minutes to recover from a 100m sprint. Some lactobacilli and streptococcus ferment lactose to lactic acid in milk. The ionization of lactic acid lowers the pH and causes denaturation of the casein (main milk protein) and other milk proteins. When this denaturation is controlled, and occurs in the right conditions, you get the yogurt or cheese.

In short, the fermentation is not used to get energy under anaerobic conditions (this misconception is very common ...). It serves to regenerate NAD+ so that glycolysis can continue to occur in the absence of O2, as glycolysis is the process that will produce ATP!

Friday, July 22, 2011

The fate of pyruvate depends on cell type and metabolic conditions. There are three main destinations for pyruvate:

(1) aerobic organisms and tissues, under aerobic conditions - pyruvate is oxidized, with loss of the carboxylic group, resulting in the acetyl group from acetyl-CoA, which is then oxidized to CO2 in the Krebs cycle;

(2) Aerobic tissues in conditions of low oxygen (muscle hypoxia, for example), some tissues under aerobic conditions (red cells, for example, because they lack mitochondria), or some anaerobic organisms - pyruvate is reduced to lactate by lactic fermentation . In muscle under conditions of hypoxia, NADH is not reoxidized to NAD+ and NAD+ is required for glycolysis. The reduction of pyruvate to lactate allows the use of as a donor of electrons to regenerate NAD+;

(3) Some tissues of plants, some invertebrates, protists and micro-organisms under anaerobic conditions or hypoxia - pyruvate is converted to ethanol + CO2 (alcoholic fermentation).

While glycolysis can occur in anaerobic conditions, this fact has a price, because it reduces the amount of ATP formed per molecule of glucose (from 30 or 32 it passes for only 2 ATP!), and, therefore, it is needed more glucose oxidation under these conditions.

What happens to pyruvate is directly related to the amount of NAD+ and FAD in the cell. As these amounts are very small, there must be mechanisms to transform the NADH + H+ and FADH2 back into NAD+ and FAD, respectively. This is done by transfer of the electrons from NADH + H+ and FADH2 to other molecules, which can occur by fermentation or respiration. The distinction between these is not (contrary to what is generally thought) that one of the processes uses directly the O2 and the other not! O2 is only required for oxidative phosphorylation, and not for the oxidation of pyruvate. Unlike aerobic metabolism that depends and are limited by the oxygen supply, the anaerobic glycolysis does not depend on the availability of oxygen and can increase speed up to 1000 times the speed at rest, ie, 2 ATP / glucose can represent many ATP/minute.

Wednesday, July 20, 2011

There is nothing new in the fact that glycolysis serves to degrade glucose, the most abundant monosaccharide in our diet. But what happens to other monosaccharides we eat? Can we degrade them for energy? If so, can we rely on glycolysis for this?
The answer to both questions is yes. Indeed, glycolysis allows the degradation not only of glucose but also of other monosaccharides such as fructose, galactose and mannose. For that, these monosaccharides have to be converted into glycolytic intermediates.FructoseFructose is obtained from the diet in foods such as fruits, honey and cereals, for example.

In extrahepatic tissues, which lack the enzyme glucocinase, fructose is converted to fructose-6-phosphate by hexokinase, the first enzyme of glycolysis. It should be noted that the hexokinase uses glucose as preferred substrate, but can also phosphorylate other monosaccharides such as fructose, for example. Since fructose-6-phosphate is an intermediate of glycolysis, it can then follow the normal glycolytic pathway.

Alternatively, in liver, fructose has to undergo a different process, because the glucocinase is very specific for glucose, failing therefore to phosphorylate fructose. In this case, fructose is phosphorylated by frutocinase, producing fructose-1-phosphate. Thereafter, the aldolase, which is the 4th enzyme of glycolysis, acts on fructose-1-phosphate, cleaving it to dihydroxyacetone phosphate and glyceraldehyde. The former molecule is already an intermediate of glycolysis, so it can proceed in that pathway, but not the latter. Therefore, the glyceraldehyde is subsequently phosphorylated by glyceraldehyde kinase, leading to glyceraldehyde-3-phosphate, which is also a glycolytic intermediate.GalactoseGalactose is found mainly in dairy products because it is a component of lactose.

Galactose is first phosphorylated to galactose-1-phosphate by galactocinase. Then galactose-1-phosphate will receive an UDP molecule, resulting in UDP-galactose. This reaction is catalyzed by galactose-1-phosphate uridyl transferase. UDP-galactose is then epimerized to UDP-glucose, by the action of UDP-galactose-4-epimerase. The UDP-glucose undergoes an exchange of its UDP portion by a phosphoryl group, producing glucose-1-phosphate, which is then converted to glucose-6-phosphate (glycolytic intermediate) by the action of phosphoglucomutase (this enzyme also participates in the metabolism of glycogen...).

Mannose
Mannose can be found, for example, in beans, peas, or similar vegetables.

Mannose is phosphorylated by hexokinase originating mannose-6-phosphate. The mannose-6-phosphate is converted to fructose-6-phosphate (a glycolytic intermediate) by fosfomanose isomerase.

The "explosion" of a grain of corn when heated is the result of the combination of three characteristics:
1. The interior of the grain (endosperm) contains, in addition to starch, about 14% water.
2. The endosperm is an excellent conductor of heat.
3. The exterior of the grain (pericarp) has great strength and rarely had flaws (cracks).
When corn is intensely heated, the water in the endosperm undergoes vaporization, creating a high pressure steam within the grain. The pericarp acts like a pressure cooker, avoiding the exit of water vapor up to a certain pressure threshold is reached. At this point, two things occur: the grain explodes, with its characteristic sound (POP!) and starchy endosperm swells abruptly, creating the soft texture.

And even now, did you know that there are no two exactly equal popcorns?

Today I leave here a link to a very interesting and fun animation on the main effects that drugs exert on the nervous system. It covers important concepts of the physiology of nerve cells, and pharmacological aspects of the drugs.

Glycolysis provides different cell molecules important for cellular proper functioning. The main donor of chemical energy in most of our processes is ATP. However, it also provides precursors for many other processes such as synthesis of amino acids, for example. Thus, it is essential to have a strict regulation of glycolysis, so that the cell can respond to different needs of ATP or other metabolites.

During his studies on fermentation of glucose by yeast, Louis Pasteur discovered that the rate and amount of glucose consumed was higher in anaerobic than in aerobic conditions! At first glance this may seem strange, because aerobic metabolism is normally associated with something more advantageous for the cell. In fact, the biochemical explanation is simple: under anaerobic conditions one molecule of glucose generates 2 molecules of ATP, but under aerobic conditions generates 30 or 32 molecules of ATP. Simplifying, if one thinks that in five minutes the yeast will need to get 30 ATP, this means that under aerobic conditions it will only need to spend a molecule of glucose in that time, while under anaerobic conditions, as the process is less profitable in the energy point of view, it is necessary to spend 15 molecules of glucose. That is, the flow of glucose through the glycolytic pathway is regulated depending on cell ATP levels (as well as adequate supplies of glycolytic intermediates to biosynthetic roles).

As I have mentioned in previous posts, glycolysis has 10 reactions, and there are three regulatory points (irreversible reactions). The enzymes that catalyze them are the hexokinase (reaction 1), PFK-1 (reaction 3) and pyruvate kinase (reaction 10). As these reactions are the limiting steps of glycolysis, changes in speed of action of their enzymes will alter the overall speed of the glycolytic pathway.

Of the three regulatory enzymes, the main one is PFK-1. This may seem strange, because indeed the most logical situation was that the main regulatory enzyme was the first ... Again, there is a very simple explanation for this. What is happening is that hexokinase is an enzyme also common to other metabolic processes (synthesis of glycogen and pentose phosphate pathway). In other words, despite being a regulatory enzyme, it is not unique to glycolysis. Thus, the main point of regulation of glycolysis has to be the second, ie, the reaction catalyzed by PFK-1.

One thing I usually tell to my students is that in this part of metabolic regulation is preferable to understand why certain molecules activate and inhibit some pathways, thus avoiding to memorize endless lists of modulators... Of course it is not always possible to make a direct argument for understand why some molecules act as activators and others as inhibitors, but whenever possible I will develop this idea... Let's move on to a list of the main modulators of glycolysis.

Activators of hexokinase:
- Fructose-1-phosphate (liver) – It competes with fructose-6-phosphate to the regulatory protein of glucocinase, canceling its inhibitory effect.
- Inorganic phosphate (Pi) – It is a player in the glycolytic process (involved in reaction 6) so it makes sense that if it has a regulatory role, is a stimulating one.Inhibitors of hexokinase:
- Glucose-6-phosphate (muscle) - It makes sense that functions as an inhibitor because it is the reaction product. If we have too much product, we will not need to continue to produce more ...
- Fructose-6-phosphate (liver) – It is the product of the following reaction (reaction 2), but can be interpreted the same way as the molecule before. That is, if we are to accumulate the intermediate formed from the reaction product, there is no point in continuing to make more product. This inhibition occurs through a protein called regulator protein of glucocinase.

Activators of PFK-1:
- Fructose-2,6-bisphosphate (liver) – It is the most significant allosteric regulator of PFK-1, reducing its affinity for the inhibitors ATP and citrate. It is produced in response to insulin and degraded in response to glucagon.
- Fructose-6-phosphate – It is the substrate, so it makes sense that if we have much substrate the enzyme is activated.
- ADP and AMP – They are produced when ATP is spent, thus indicating a low energy state. Therefore, it makes perfect sense that they can activate glycolysis, so that the cell can replenish their normal energy values. They activate the enzyme because they relieve the inhibition caused by ATP.Inhibitors of PFK-1:
- Glucagon (liver) - This hormone is produced in a state of hypoglycemia and aims to raise the concentration of glucose in the blood. So it makes perfect sense that it inhibits glycolysis, because this process consumes glucose, which will further accentuate the reduced blood glucose concentration. As mentioned earlier, the glucagon decreases the levels of fructose-2,6-bisphosphate
- ATP - The main objective of glycolysis is to produce energy (ATP). So if the cell already has ATP, it makes sense that glycolysis is inhibited, thus preventing an unnecessary waste of a precious metabolic fuel as glucose! ATP inhibits PFK-1 because it decreases the affinity of the enzyme for its substrate, fructose-6-phosphate.
- Citrate – It stresses the inhibitory effect of ATP. This molecule is the first intermediate of the following step of aerobic catabolism, the Krebs cycle. So if we are accumulating Krebs cycle intermediates, it is useless to continue to perform glycolysis.
- Phosphoenolpyruvate – It is an intermediate of glycolysis that is formed in the penultimate reaction. If there is an accumulation of this intermediate, the reactions above have to be inhibited in order to prevent a further accumulation of the molecule.- H+ - This enzyme is particularly sensitive to changes in pH, functioning as a "switch" that turns off, for example, when we make an exaggerated lactic fermentation (produces H+), preventing an even greater acidification.

Activators of pyruvate kinase:
- ADP - The reason is the same as mentioned above for the PFK-1, ie, is an indicator of an energy deficit, so it will lead to an activation of glycolysis.
- Fructose 1,6-bisphosphate - an intermediate of glycolysis that is formed in a reaction prior to the one catalyzed by pyruvate kinase. So if we are accumulating an intermediate produced in an earlier stage, we have to activate this enzyme in order to counteract this accumulation (as when a dam is accumulating too much water, and to restore normal values ​​is necessary to open the gate ...).
- Dephosphorylation (liver) - Induced, for example, by insulin, which makes sense, given that insulin is produced in a situation of excess blood sugar (hyperglycemia) and will activate the process (one of them is glycolysis!) that consume glucose in order to lower the blood glucose concentration.Inhibitors of pyruvate kinase:
- ATP – It is a carrier of chemical energy and one of the end products of glycolysis, so there is no need to continue the breakdown of glucose. It decreases the affinity of the enzyme for phosphoenolpyruvate.
- Acetyl-CoA – It is the molecule in which the product of this reaction (pyruvate) is converted in the case of aerobic catabolism. Therefore, if acetyl-CoA accumulates, it makes no sense to continue to synthesize pyruvate, so the enzyme is inhibited.
- Long-chain fatty acids.
- Phosphorylation (liver) - Induced, for example, by the action of glucagon, which, as mentioned earlier, will have as main function to raise blood glucose levels. To this end, it inhibits, for example, glycolysis.
- NADH - as we shall see in more detail when I speak of cellular respiration, NADH has potential to create molecules of ATP, which signals a high energy state of the cell. In this situation, it is not necessary to resort to glycolysis for more energy.
- Alanine - This amino acid (one of the 20 standard amino acids) can lead to pyruvate (the reaction product of pyruvate kinase!), by removal of its amino group. So if there is a molecule that can directly lead to pyruvate, we do not need to spend more glucose.

In short, we can make some generalizations about the regulation of metabolic pathways, which will be useful to understand the regulation of other processes.
First, energy molecules such as ATP, or potential energy, such as NADH are, in general, inhibitors of catabolism. This is very easy to understand if we think that the main objective of the catabolism is to produce energy. If the cell already has this ability it does not need to degrade more nutrients to produce energy! The oppposite reasoning applies to ADP, AMP and NAD+, because any one of these molecules indicates an energy deficit in the cell (remember that when we spend ATP we obtain ADP or AMP, and when we spend NADH we obtain NAD+...) so it will be necessary to restore energy levels, and this activates the catabolism.
Second, the reaction product or intermediates formed from this (products of reactions following the reaction we are considering) are inhibitors. On the other hand, the substrate, or intermediaries which will originate the substrate (formed in reactions prior to the reaction that we are considering) are activators.

Sunday, July 17, 2011

The biosynthesis of proteins was also a topic chosen by Dr. Baum for one of his biochemical songs. In this case, using the known My Bonnie Lies Over the Ocean as a starting point. Here is the link to download the music.

Saturday, July 16, 2011

Making a metabolic map with just some pathways is no an easy task. With all the metabolic pathways, it is significantly worse. The link that I leave in this post lets you see the most complete metabolic map that I know. It is worthwhile to see him, even if only to have biochemical nightmares in the following night... :)

Our body has a remarkable metabolic flexibility! Consider, for example, that we can adapt to situations as opposed as being 8-9 hours without eating (when you sleep, for example), or eat a very caloric meal.

Or else do a very intense workout in a short space of time, or a more moderate and longer, or simply stay at home. This ability to properly handle these opposites is a consequence of a strict regulation of our metabolic pathways.

The regulation of metabolic processes is, in my opinion the main point for a correct understanding of metabolism.

Before I start talking about specific regulation of each pathway, it is important to address some more general concepts ...
First, what is the regulation of metabolic pathways? It is the process by which the overall speed of each process is changed. Please note that when it comes to regulation, it does not mean necessarily inhibition, because the metabolic pathways can be activated or inhibited.
All pathways have at least one specific reaction of this process, which is irreversible. This ensures the cell two important ways:
1. Causes metabolic pathways do not occur in both directions, as a consequence only of the mass flow. That is, if a pathway produces the molecule X and the cell needs to produce more X, it is not because there is already this molecule within the cell that the same will be degraded.
2. They are used to regulate a specific metabolic pathway without affecting other processes, namely, the opposite process. To understand this we can think of two opposing processes, glycolysis (breakdown of glucose) and gluconeogenesis (glucose synthesis), for example. In cells the two processes do not occur simultaneously because there was no point being to degrade and synthesize glucose at the same time. Therefore, when one is active, the other must be inhibited. If both were catalyzed by the same enzymes, it was impossible to activate a process and inhibit another. Or both were activated or inhibited... How do we get around this problem? Using at least one enzyme specific for each process! So if I have a specific enzyme in glycolysis (there are 3 in fact ...) that does not act in gluconeogenesis, I can activate or inhibit this process without affecting the opposite. J

It is these very specific and irreversible reactions that are catalyzed by enzymes called regulatory. The regulatory enzymes are enzymes that act as a kind of metabolic pathways valves, allowing "flow" more intermediaries, if you lack more product, or accumulate these intermediates, if there is enough product. The reactions catalyzed by these enzymes are often referred to as points of regulation, considering that they are the limiting steps (slower) of the process to which they belong. So if their speed is increased, the overall speed of the pathway where they are located increases, and if its speed is decreased, the overall speed of the process decreases.

There are four types of regulation of metabolic pathways:1. Substrate availability - is the fastest method of regulation and affects all the enzymes in each pathway. Basically, if there is few substrate, the enzymes will not be able to act at its maximum speed, and if there is no substrate, the enzymes stop.2. Allosteric regulation - is the fastest way to regulate only certain specific enzymes, called regulatory enzymes. This form of regulation requires the presence of molecules (allosteric modulators) that will interact with the enzymes, leading to structural changes that can make the enzyme faster or slower (positive and negative modulators, respectively).

3. Hormonal regulation - is a slower process than the allosteric regulation, and involves the production of hormones in response to a stimulus. Hormones are released into the bloodstream and will act on target cells. Usually its action culminates in the phosphorylation or dephosphorylation of regulatory enzymes, altering their catalytic efficiency (activated or inhibited, depending on the enzyme in question). This effect is called reversible covalent modification.

4. Changes in the concentration of enzymes - This is the slowest way of regulation and include changes in rates of synthesis and degradation of enzymes, altering their concentration. For example, if a cell wants to activate a metabolic pathway, it can do so by increasing the amount of the enzymes of this pathway. Since the substrate is not limiting, the overall rate of conversion of substrate to product will increase. The opposite effect is observed by the opposite reasoning.